The continuing overall objective of the proposed research is to determine the various mechanisms by which the amount and activity of the specific non-secreted protein with a known function is regulated in uterine tissue by ovarian hormones. The uterine protein to be studied is the enzyme glucose-6-phosphate dehydrogenase (G6PD) which is inducible by estradiol and which functions as the first enzyme of the pentose phosphate pathway. SPECIFIC AIMS: 1) To determine the mechanisms by which the amount of the mRNA for G6PD in uterine tissue is regulated by estradiol, 2) to determine mechanisms by which the rate of translation of the mRNA for G6PD in uterine tissue is regulated by estradiol and the cofactor of the enzyme, NADP+, 3) to characterize a precursor form of G6PD (pre-G6PD), determine its possible "functional" roles in the uterus other than to serve as a precursor for cytosolic G6PD and to determine the estrogen dependent mechanism by which pre-G6PD is converted to G6PD, and 4) to determine the mechanisms of inactivation and then degradation of uterine G6PD and the manner by which the inactivation or degradative process is inhibited by estradiol. A cDNA to uterus mRNAG6PD will be prepared and used to directly measure the rates of synthesis, processing and degradation of mRNAG6PD in the uterus. Uterine tRNA from estrogen-induced ovariectomized mature rats will be evaluated for its ability to enahance the synthesis of a "translation restriction zone" in mRNAG6PD. Trypsin mapping techniques and changes in molecular weights of pre-G6PD and G6PD and G6PD will be used to characterize pre-GSPD. A G6PD inactivating activity in the uterus cytosol will be purified and characterized as a potential estrogen regulated first step in the degradation of the uterine G6PD. The study of mechanisms involved in regulation of this enzyme in the uterus is important because of the key role this enzyme plays in general uterine metabolism and because the regulation of this enzyme will likely serve as a model system to provide techniques and a basis for the study of the mechanisms involved in the regulation of other functional, non-secreted uterine proteins.